ca-circSCN8A promotes HPASMCs ferroptosis via LLPS initiated R-Loop

Li, M.; Hao, Y.; Song, X.; Liu, H.; Zhang, C.; Zhang, J.; Sun, H.; Zheng, X.; Zhang, L.; Yu, H.; Ma, C.; Zhao, X.; Zhu, D.

doi:10.1101/2025.01.21.634205

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bioRxiv · 10.1101/2025.01.21.634205

ca-circSCN8A promotes HPASMCs ferroptosis via LLPS initiated R-Loop

Li, M.·Hao, Y.·Song, X.·Liu, H.·Zhang, C.·Zhang, J.·Sun, H.·Zheng, X.·Zhang, L.·Yu, H.·Ma, C.·Zhao, X.·Zhu, D.

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Abstract

BACKGROUNDFerroptosis has been implicated in pulmonary hypertension. Chromatin-associated RNAs are linked to ferroptosis. However, their role in pulmonary vascular ferroptosis in PH is unexplored. METHODSBioinformatics, Sanger sequencing, RNase R and others were employed to identify differentially higher expression of ca-circSCN8A. Functional gain and loss assays were employed to unveil the role of ca-circSCN8A in hypoxia-induced redox-dependent ferroptosis in HPASMCs and PH mice model. Interaction between ca-circSCN8A and FUS was detected via RNA immunoprecipitation and pull-down assays. FRAP, CHIRP-qPCR, MDA, GSH and GSSG were conducted to explore the potential molecular mechanism. RESULTSca-circSCN8A was firstly identified and confirmed to be upregulated in PH. Upregulation of ca-circSCN8A can promote hypoxia induced ferroptosis in HPASMCs. Under hypoxic conditions, ca-circSCN8A promoted the lactylation of FUS by recruiting EP300, leading to formation of LLPS of ca-circSCN8A/FUS/EP300. LLPS maintained the stability of the R-Loop formed by ca-circSCN8A and ferroptosis related-gene SLC7A11 promoter that inhibit the transcription of the SLC7A11 gene, further result in the disruption of the redox balance and causes ferroptosis in HPASMCs. CONCLUSIONSca-circSCN8A recruits EP300 to promote the lactylation of FUS, thereby promoting LLPS formation of ca-circSCN8A/EP300/FUS. Further, the LLPS motivates ca-circSCN8A to form R-loop with its non-host SLC7A11 and participating in the regulation of hypoxic induced HPASMCs redox-dependent ferroptosis. This is the first confirmation that circRNAs forming R-loops with non-host genes regulated by LLPS. Our findings indicated that ca-circSCN8A plays an important role in ferroptosis of hypoxic induced HPASMCs and may be a potential target for the treatment of PH.

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Li, M., Hao, Y., Song, X., Liu, H., Zhang, C., Zhang, J., Sun, H., Zheng, X., Zhang, L., Yu, H., Ma, C., Zhao, X., Zhu, D.. 2025-01-22. ca-circSCN8A promotes HPASMCs ferroptosis via LLPS initiated R-Loop. https://doi.org/10.1101/2025.01.21.634205

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